Effect Of Trichostatin A On Cystoskeleton,Cycle And Histone Acetylation Of Donor Cells

1.It has been widely suggested that poor ability of oocyte cytoplasmic factors to globally erase the epigenetic modifications of a somatic cell is the major reason of the inefficiency of nuclear transfer success.Trichostatin A(TSA)is a histone deacetylase inhibitor.In this study, bovine fetal fibroblasts passaged 5 times were seeded into DMEM plus 10% FBS containing 75nmol/L of TSA for 6,12,24h.These levels were selected to produce time-dependent effects. Cells were subsequently subjected to essay for histone acetylation of H3K18 levels by confocal microscope.The effect of Trichostatin A treatment donor cells on in vitro development of bovine nuclear transfer (NT) embryos that constructed with bovine fetal fibroblast cells and cell cycle was investigated.these resuits show that TSA can increase the levels of histone acetylation of H3K18 after bovine fetal fibroblast Cells treated with 75nmol/L TSA for 12h or 24h( p<0.05);75nmol/L TSA treatment of donor cells for 12h increased blastocyst development compared to controls (23.5%vs15.7%,p<0.05).Conclusion: TSA enhances the reprogramming in terms of high histone acetylation of doner cells.2.We examined global changes in the acetylation of histones in bovine oocytes during meiosis. Trichostatin A is a specific inhibitor of histonedeacetylase(HDAC).Immunocytochemistry with specific antibodies against acetylated lysine residues on histones H3 showed that acetylation of H3K18 decreased to undetectable or negligible levels in the oocytes during meiosis;Intense fluorescence signals for acetylated H3K18 were detected when meiosis oocytes treated with TSA;the signals reappeared 3h after transfer to free TSA medium.our results suggest that HDAC is able to deacetylate histones during meiosis,HAT is inactive during meiosis.3.Basically,no alteration was observed in cell morphology after treatment with TSA,some cells treated with TSA appeared flattened.Fluorescence-activated cell sorting analysis was used to assess the cell cycle-stage distribution,60.1% of TSA-treated cells were at the G0/G1 phase of the cell cycle,marked difference existed in the G0/G1 and S stage rate of treated group and control group( p<0.05).Cells were subsequently subjected to essay for -tubulin by confocal microscope.the resuits show that there is a clear production in tubulin levels in TSA-treated cells, it proved that TSA-treated cells is active in another facts.